1. Field of the Invention
This invention generally relates to the use of nucleic acid sequences encoding growth factors to promote angiogenesis and wound healing. In particular, described herein are methods and compositions for enhancing flap and skin graft survival by administering a nucleic acid sequence encoding hepatocyte growth factor.
2. Background of the Invention
Hepatocyte growth factor (HGF) functions as a growth factor for particular tissues and cell types. HGF was initially identified as a mitogen for hepatocytes. Michalopoulos et al., Cancer Res., 44:4414-4419 (1984); Russel et al., J. Cell. Physiol., 119:183-192 (1984); Nakamura et al., Biochem. Biophys. Res. Comm., 122:1450-1459 (1984). Nakamura et al. (supra), reported the purification of HGF from the serum of partially hepatectomized rats. Subsequently, the subunit structure of HGF was determined when HGF was purified from rat platelets. Nakamura et al., Proc. Natl. Acad. Sci. USA, 83:6489-6493 (1986); Nakamura et al., FEBS Letters, 224:311-316 (1987). Human HGF (“huHGF”) has also been purified from human plasma. Gohda et al., J. Clin. Invest., 81:414-419 (1988).
Comparisons of the amino acid sequence of rat HGF and huHGF revealed that the two sequences are highly conserved and have the same characteristic structural features. For example, the length of the four kringle domains in rat HGF is exactly the same as in huHGF, and the location of cysteine residues are in exactly the same positions. This is an indication that the three-dimensional structure of the two proteins is similar. Okajima et al. Eur. J. Bioch., 193:375-81 (1990); Tashiro et al., Proc. Natl. Acad. Sci., USA, 87:3200-4 (1990).
Furthermore, several reports revealed close sequence homology between HGF and scatter factor (SF). Gherardi and Stoker, Nature, 346:228 (1990); Weidner et al., J. Cell Biol., 111:2097-2108 (1990); Coffer et al., Biochem J., 278:35-41 (1991). SF is a polypeptide that stimulates dissociation of epithelial cell colonies in monolayer culture. Gherardi et al., Proc. Natl. Acad. Sci. USA, 86:5844-5848 (1989). In fact, there now is evidence indicating that the two factors are identical; they are identical in structure and biological activity. Weidner et al., Proc. Natl. Acad. Sci. USA, 88:7001-5 (1991); Bhargava et al., Cell Growth Differ. 3:11-20 (1992); Naldini et al., EMBO J., 10:2867-78 (1991); Furlong et al., J. Cell Sci., 100:173-7 (1991). HGF and HGF variants are described further in U.S. Pat. Nos. 5,227,158, 5,316,921, and 5,328,837.
Binding of HGF to its receptor is believed to be conveyed by a functional domain located in the N-terminal portion of the HGF molecule. Matsumoto et al., Biochem. Biophys. Res. Commun., 181:691-699 (1991); Hartmann et al., Proc. Natl. Acad. Sci. USA., 89:11574-11578 (1992); Lokker et al., EMBO J., 11:2503-2510 (1992); Lokker and Godowski, J. Biol. Chem., 268:17145-17150 (1991). The HGF receptor is usually referred to as “c-Met” or “p190MET” and typically comprises, in its native form, a 190-kDa heterodimeric (a disulfide-linked 50-kDa α-chain and a 145-kDa β-chain) membrane-spanning tyrosine kinase protein. Park et al., Proc. Natl. Acad. Sci. USA, 84:6379-6383 (1987). The c-Met protein becomes phosphorylated on tyrosine residues of the 145-kDa β-subunit upon HGF binding.
Various biological activities have been described for HGF and its receptor. See, generally, Chan et al., HEPATOCYTE GROWTH FACTOR—SCATTER FACTOR (HGF-SF) AND THE C-MET RECEPTOR, Goldberg and Rosen, eds., Birkhauser Verlag-Basel (1993), pp. 67-79). For example, HGF has been shown to be a mitogen for certain cell types, including melanocytes, renal tubular cells, keratinocytes, certain endothelial cells and cells of epithelial origin. Matsumoto et al., Biochem. Biophys. Res. Commun., 176:45-51 (1991); Igawa et al., Biochem. Biophys. Res. Commun., 174:831-838 (1991); Han et al., Biochem., 30:9768-9780 (1991); Rubin et al., Proc. Natl. Acad. Sci. USA, 88:415-419 (1991). HGF has also been described as an epithelial morphogen, Montesano et al., Cell, 67:901-908 (1991), and therefore, HGF has been postulated to be important in tumor invasion, Comoglio, HEPATOCYTE GROWTH FACTOR—SCATTER FACTOR (HGF-SF) AND THE C-MET RECEPTOR, Goldberg and Rosen, eds., Birkhauser Verlag-Basel (1993), pp. 131-165. Until now, the intramuscular delivery of an HGF gene to promote flap and skin graft survival has not been described.
The use of skin flaps has gained increased acceptance and use in the course of reconstructive surgery, as well as in other forms of surgery. However, these techniques continue to be plagued by problems having to do with survival of the skin flaps which is, at least in part, due to the inefficient revascularization at the surgical site. Indeed, a number of approaches have been considered or evaluated for improving skin flap survival. See, for example, Waters et al., which provides a comparative analysis of the ability of five classes of pharmacological agents to augment skin flap survival in various models and species. Annals of Plastic Surgery, 23(2):117-22 (1989). Nevertheless, there still remains a need in the art for compositions and methods for enhancing survival of flap and skin grafts.